S-Hydroxynitrile lyase catalyzes the reaction between hydrocyanic acid and aldehyde or ketone to generate optically active cyanohydrins. Optically active cyanohydrins are important intermediates for synthesizing medicines. Accordingly, it can be said that S-hydroxynitrile lyase is an industrially important enzyme.
S-hydroxynitrile lyases derived from Cassava (Manihot esculenta), Pará rubber tree (Hevea brasiliensis), and poaceous plants (i.e., sorghum (Sorghum bicolor)) have been known. Industry may prefer recombinant S-hydroxynitrile lyase in addition to wild-type enzymes, because separation of enzymes from organisms incurs a high cost.
Production of recombinant S-hydroxynitrile lyase requires a step of separating an enzyme from a solution of disrupted host cells without loss in its activity. A technique that employs chromatography such as ion-exchange chromatography is the most common method for separating enzymes. Due to the high cost thereof, however, a separation technique via heating is preferable at industrial levels.
Wagner et al. analyzed the crystal structure of Hevea brasiliensis-derived S-hydroxynitrile lyase and reported that this enzyme belongs to the α/β hydrolase superfamily. According to this report, the catalytic triad of Ser80, His235, and Asp207 constitute the enzyme's active center. It is located deep inside the protein, and it is linked to the outside through a narrow hydrophobic channel (Wagner U G. et al., Structure, 1996, Jul. 15, 4(7), pp. 811-822). In order to improve substrate receptivity of enzymes, modified S-hydroxynitrile lyase was developed. In this S-hydroxynitrile lyase, bulky amino acids are substituted with smaller amino acids, in the hydrophobic channel. (JP Patent Publication (Kokai) No. 2000/125886 A). However, modification of S-hydroxynitrile lyase for other purposes than the substrate receptivity, for example, modification to improve thermostability, has not yet been reported.